c-Met, a member of the tyrosine kinase superfamily, is the receptor for Hepatocyte Growth Factor (HGF). Binding of HGF to c-Met leads to receptor dimerization or multimerization, phosphorylation of multiple tyrosine residues in the intracellular region, catalytic activation, and downstream signaling. c-Met is also activated via ligand-independent mechanisms, including receptor over-expression, amplification, and mutation. c-Met activation enhances cellular proliferation, migration, morphogenesis, survival (including protection from apoptosis), and protease synthesis, characteristics that are associated with invasive cell phenotype and poor clinical outcomes and drug resistance in cancer patients. The c-Met signaling pathway is one of the most frequently dysregulated pathways in human cancers, and occurs in virtually all types of solid tumors.
c-Met is over-expressed in many different type of cancer cells and plays important role in mesenchymal-epithelial interaction and embryogenesis (X Liu et al, Trends in Molecular Medicine, 2009; E M Rosen et al, JCB, 1994; C Birchmeier et al, Trends in Cell Biology, 1998). It is also associated with drug resistance in lung cancer (J Engelman et al, Science, 2007). This makes it a useful target for therapeutic and diagnostic applications.
In addition to many small molecular inhibitors that have been studied for therapeutic effects, such as PHA-665752, SU11274, and ARQ197 (N Puri et al, Cancer Research, 2007; S Berthou et al, Oncogene, 2004; T Underiner et al, Anti-Cancer Agents in Medicinal Chemistry, 2010), there are also anti-MET antibodies that have been developed. Examples of anti-MET antibodies include the mouse monoclonal antibody DN-30 and the humanized one-armed monoclonal antibody onartuzumab. These antibodies have been evaluated for PET imaging applications (L R Perk et al, Eur J Nucl Med Mol Imaging, 2008; E M Jagoda et al, JNM, 2012).
There exists a need for antagonist antibodies to human c-Met, binding of which to the α-chain of human c-Met facilitates internalization of the receptor from the cell surface. There is also a need for antagonist antibodies to human c-Met, which binding to the α-chain of human c-Met facilitates internalization of the receptor from the cell surface in cells comprising c-Met variants containing gain of function mutations. There is also a need for antagonist antibodies to human c-Met which induce c-Met degradation and reduction of phosphorylated c-Met. Such antagonist activities could decrease the number of available binding sites for HGF on tumor cell surfaces, and terminate the pathway activation caused by overexpression, amplification, or mutation of c-Met. At the same time, such antagonist antibodies should inhibit HGF binding to c-Met and HGF-induced c-Met activation, and induce little or no agonist activity themselves.
The novel selection and characterization of several novel human scFv antibodies against human c-MET from a naive human scFv phage display library is described herein. scFv clones were confirmed to bind to cell surface target, and reformatted into cys-diabodies. Cys-diabodies have been successfully expressed in bacteria, cell lines, and tumor models and tested for affinity. The low immunogenicity, high affinity and varied pharmacokinetic characteristics of these fully human antibody fragments give them great potential in both in vivo and in vitro applications. Furthermore, the good affinity and low immunogenicity of these antibodies make them very useful for both in vivo and in vitro applications. These novel human antibodies can greatly help the study of drug resistance in EGFR targeted therapies, and improve the diagnosis and treatment of cancer patients.